Chromosomal duplication, an event tightly coupled to cell division, is a fundamental biological process of every organism. Like many biochemical pathways, this event is regulated at the first step, the initiation of a cycle of chromosomal replication. The long range objective of this proposed research is to understand in biochemical terms, the mechanism of initiation of Escherichia coli chromosomal replication, its regulation, and how these events are coupled to other cellular processes. The specific aims of the proposed research are to focus on the mechanism of suppression of conditionally defective dnaA gene product, a protein required for initiation of E. coli chromosomal replication, by extragenic suppressors of dnaA by using a combined approach of molecular biology and enzymology. This study will provide insight into 1). other replication enzymes which interact with dnaA protein in the initiation of chromosomal replication, 2). the series of events involved in this process, and 3). alternative mechanisms of chromosomal duplication which occur by initiating replication at sites other than at the chromosomal origin. A biochemical understanding of the initiation process which is so tightly coupled to cell division, how it is regulated, and alternative pathways of chromosomal duplication will further our understanding of how these processes may occur in higher organisms, what may cause malignant growth of normally quiescent cells, and eventually how this aberrant process can be controlled or prevented.